Pulmonary arterial hypertension (PAH) is a life-threatening disease characterized by abnormally elevated pulmonary pressures and right heart failure. While the etiology of PAH is unknown, progressive loss and impaired regeneration of microvessels suggest that defective angiogenesis may be a major contributor to PAH pathogenesis. Control of angiogenesis is dependent on proper activation of the vascular endothelial growth factor (VEGF) pathway and while some studies show evidence of elevated VEGF pathway components within PAH vascular lesions, others demonstrate that pulmonary microvascular endothelial cells (PMVECs) from PAH patients form small and disorganized vascular networks under VEGF stimulation. Our inability to explain these observations is due in large part to an incomplete understanding of how response to VEGF signaling is regulated in PMVECs. We have discovered a novel link between VEGF and Wnt signaling where endothelial-derived Wnt7a, a Wnt ligand, is required to fully activate VEGF signaling in PMVECs via ROR2, a receptor belonging to the Wnt/PCP pathway. Failure of PAH PMVECs to respond to VEGF correlates with reduced Wnt7a production and is corrected by exogenous Wnt7a. Based on these studies, we propose that reduced angiogenesis in PAH is caused by lack of crosstalk between the VEGF and Wnt7a/ROR2 pathways, which prevents PMVECs from assuming a tip cell phenotype to direct vessel assembly. To establish this, we will apply analytical protein analysis and gene editing techniques to establish how VEGFR2- ROR2 receptor crosstalk modulates VEGF response in PMVECs (Aim 1) and will use live imaging 3D angiogenesis to study tip cell formation by PMVECs in response to VEGF and Wnt7a (Aim 2). For Aim 3, we will use transgenic murine models to demonstrate that conditional deletion of Wnt7a and ROR2 levels in PMVECs can worsen hypoxia-induced small vessel loss and hemodynamics; we will also begin testing a novel nanoparticle-based therapeutic to determine whether restoring Wnt7a expression can improve these parameters in Sugen/hypoxia rats. Addressing how VEGF and Wnt orchestrate pulmonary angiogenesis will provide not only fundamental mechanistic insights into the regulation of angiogenesis but facilitate development of Wnt7a-based therapeutics that could prevent small vessel loss in PAH.